Efficacy of netropsin dihydrochloride against the viability, cytopathogenicity and hemolytic activity of Trichomonas vaginalis clinical isolates.

Trichomonas vaginalis (T. vaginalis) is a common sexually transmitted infection, affecting the urogenital tract. Trichomoniasis is customarily treated with metronidazole (MTZ). MTZ is known to cause undesirable side effects and there is several reports on MTZ resistant T. vaginalis. Thus, the present study aimed to in-vitro evaluate the activity of DNA minor groove binder drug ''Netropsin dihydrochloride'' against metronidazole-sensitive T. vaginalis isolates (G and U isolates) and resistant T. vaginalis isolate (ATCC50138) (R isolate). Netropsin was tested at concentrations ranging from 3.5 to 200 µg/ml. It showed effectiveness against all isolates with MLC of 12.5 µg/ml for G and U isolates and of 25 µg/ml for R isolate. Cytotoxicity assay of isolates exposed to the respective MLC of netropsin for 42 h showed a highly significant reduction in the death percentage of MCDK cell line as compared to the effect elicited by drug free controls. The hemolytic activity was evaluated by hemolytic assay and by monitoring the interaction of T. vaginalis isolates with human erythrocytes by inverted microscopy and scanning electron microscopy. The hemolytic assay showed (0%) hemolysis of RBCs incubated with T. vaginalis isolates treated with the corresponding MLC of netropsin for 24 h. Scanning electron microscopy revealed cytoskeletal deformities of netropsin treated isolates. Taken together, these observations suggest that netropsin is a promising therapy for T. vaginalis infection affecting its viability, virulence, cytopathogenic and hemolytic activity with a mechanism of action that might overcome T. vaginalis resistance to metronidazole.

Efficient therapeutic effect of Nigella sativa aqueous extract and chitosan nanoparticles against experimentally induced Acanthamoeba keratitis.

Acanthamoeba keratitis (AK) is a devastating, painful corneal infection, which may lead to loss of vision. The development of resistance and failure of the currently used drugs represent a therapeutic predicament. Thus, novel therapies with lethal effects on resistant Acanthamoeba are necessary to combat AK. In the present study, the curative effect of Nigella sativa aqueous extract (N. sativa) and chitosan nanoparticles (nCs) and both agents combined were assessed in experimentally induced AK. All inoculated corneas developed varying grades of AK. The study medications were applied on the 5th day postinoculation and were evaluated by clinical examination of the cornea and cultivation of corneal scraps. On the 10th day posttreatment, a 100% cure of AK was obtained with nCs (100 µg/ml) in grades 1 and 2 of corneal opacity as well as with N. sativa 60 mg/ml-nCs 100 µg/ml in grades 1, 2, and 3 of corneal opacity, highlighting a possible synergistic effect. On the 15th day posttreatment, a 100% cure was reached with N. sativa aqueous extract (60 mg/ml). Moreover, on the 20th day posttreatment, N. sativa (30 mg/ml) provided a cure rate of 87.5%, while nCs (50 µg/ml) as well as N. sativa 30 mg/ml-nCs 50 µg/ml yielded a cure rate of 75%; the lowest percentage of cure (25%) was obtained with chlorhexidine (0.02%), showing a non-significant difference compared to the parasite control. The clinical outcomes were in agreement with the results of corneal scrap cultivation. The results of the present study demonstrate the effectiveness of N. sativa aqueous extract and nCs (singly or combined) when used against AK, and these agents show potential for the development of new, effective, and safe therapeutic alternatives.

Cytotoxic effect of organic solvents and surfactant agents on Acanthamoeba castellanii cysts.

Acanthamoeba castellanii is a protozoan parasite that may cause sight-threatening keratitis in some individuals. Its eradication is difficult because the trophozoites encyst making organisms highly resistant to anti-amoebic drugs. To test new anti-Acanthamoeba agents, usually having low water solubility, organic solvents and surfactant agents should be used. Therefore, the lethal effect of different concentrations of the solvents acetone, methanol, ethanol, and DMSO and surfactant agents Tween 20, Tween 80, and Triton X-100 was tested. The minimal inhibitory concentrations (MIC) were determined against Acanthamoeba cysts. Results of the present study showed that the MIC for ethanol, methanol, acetone and DMSO was 25, 12.5, 12.5, and 10%, respectively and for Tween 20, Tween 80, and Triton X-100 was 0.25, 0.06, and 0.03%, respectively. There was no significant inhibitory effect on the multiplication of Acanthamoeba cysts as compared to parasite control when using the concentrations 3.12% for ethanol, 1.6% for methanol and acetone, 1.25% for DMSO, and 0.016% for Tween 20. On the other hand, both Tween 80 and Triton X-100 showed highly significant difference in comparison to parasite control almost among all the range of concentrations used in this study, and both showed lethal effect of 19 and 27.2%, respectively at their least concentration.

The impact of vinegar on pathogenic Acanthamoeba astronyxis isolate.

Acanthamoeba keratitis (AK) is a severe corneal disease that was reported by WHO as the second most common infectious cause of blindness after trachoma; contact lens wear is considered one of the main risk factors in its transmission. Thus, the treatment of AK is crucial but, the inability of medical agents to completely eradicate the resistant cyst, together with their toxic effects, suggest that new agents are needed. Vinegar has been known long ago as a simple and available disinfectant with antimicrobial effects, so the present study aimed to test the effect of different concentrations of vinegar solution on Acanthamoeba astronyxis isolate, along the period of 1 h in comparison to parasite and chlorhexidine controls. Post hoc test analysis revealed a highly significant difference between the vinegar-treated parasites and both controls, as regards the viable and non-viable mean cysts count. Vinegar concentration of 5% exhibited the highest mean of non-viable cysts along the time intervals, while the lowest was shown with 0.04% where also, no viable cysts were detected at 60 min. All tested concentrations behaved in a time-dependent manner. There was a positive correlation with a significant outcome between the different concentrations and the mean of the non-viable parasites along time. Transmission electron microscopy of treated cysts revealed corrugated altered cell wall with loss of ridges and detachment and shrinkage of content. Treated trophozoites showed flattening of the acanthopodia with thinned out plasma membrane and degenerated cytoplasmic content. The study highlighted the potential use of vinegar as an adjuvant in the prevention and treatment of AK.

In vitro anti-Trichomonas vaginalis activity of Pistacia lentiscus mastic and Ocimum basilicum essential oil.

Trichomonas vaginalis is a protozoan parasite that causes trichomoniasis; a cosmopolitan sexually transmitted disease. Metronidazole is the drug of choice for T. vaginalis infections. The increase in metronidazole resistant parasites and undesirable side effects of this drug makes the search for an alternative a priority for the management of trichomoniasis. Pistacia lentiscus mastic and Ocimum basilicum oil are known for their antibacterial, antifungal, antiviral and antiprotozoal effects. The present study was carried out to investigate the in vitro effects of P. lentiscus mastic and O. basilicum oil on T. vaginalis trophozoites. The effects of different concentrations of P. lentiscus mastic (15, 10 and 5 mg/ml) and different concentrations of O. basilicum oil (30, 20 and 10 µg/ml) on multiplication of trophozoites at different time points (after 24, 48, 72 and 96 h) were determined, also morphological changes were reported by transmission electron microscopy (TEM). The results showed that both plants caused an inhibition of growth of T. vaginalis trophozoites. The minimal lethal concentration of P. lentiscus mastic was 15 mg/ml after 24 h incubation, 10 mg/ml after 48 h and 5 mg/ml after 96 h. The minimal lethal concentration of O. basilicum oil was 30 µg/ml after 24 h incubation, 20 µg/ml after 48 h and 10 µg/ml after 96 h. TEM study of trophozoites treated by P. lentiscus mastic or by O. basilicum oil showed considerable damage of the membrane system of the trophozoites, and extensive vacuolization of the cytoplasm. These results highly suggest that P. lentiscus mastic and O. basilicum oil may be promising phytotherapeutic agents for trichomoniasis treatment.

A comparative study between excretory/secretory and autoclaved vaccines against RH strain of Toxoplasma gondii in murine models.

Toxoplasma gondii is an obligate intracellular protozoan that has a major importance in public health, in addition to veterinary medicine. Therefore, the development of an effective vaccine for controlling toxoplasmosis is an important goal. Excretory/secretory antigens (ESA), were previously identified as potential vaccine candidates, proved to play important roles in the pathogenesis and immune escape of the parasite. In addition, autoclaved Toxoplasma vaccine (ATV) is a special type of killed vaccine, recently characterized. The aim of the present work was, to compare between excretory/secretory and ATV against RH strain of T. gondii in mice based on; parasitological and histopathological levels. Tachyzoites were harvested from peritoneal exudates of infected mice and were used for challenge infection and vaccine preparation. BCG was used as an adjuvant. Mice were allocated equally into five groups; they were vaccinated intradermally over the sternum. The results of this study showed that the survival time after challenge, extended up to 16 days in ESA vaccinated group and up to 15 days in autoclaved Toxoplasma vaccinated group. ESA vaccinated group exhibited a profound decrease in parasite load following parasite challenge with a higher percentage of reduction in parasite count in all examined organs than the autoclaved Toxoplasma vaccinated group. The histopathological picture of the liver in both immunized groups, revealed marked reduction in the pathological changes observed as compared to controls, especially in ESA vaccinated group. It was concluded that vaccination with ESA showed more promising results versus ATV, as demonstrated by the survival rate of vaccinated mice, tachyzoites count and histopathological examination.